UV radiation is the leading environmental cause of skin cancers. However, despite the known etiological effects of UV exposure, methods for detecting initial UV-DNA damage and the mutational processes that follow are severely limited. There is an urgent need for a high-throughput method that can directly detect UV-DNA damage at the genome level in human cells. Here, we propose to develop a new method based on damaged DNA immunoprecipitation (DDIP) enrichment followed by single- molecule, real-time (SMRT) DNA sequencing. The development of DDIP-SMRT seq will have a major impact on our ability to directly assess the effects of UV-radiation and other environmental DNA damaging agents on cellular DNA. To develop DDIP-SMRT seq, we propose two aims. In aim 1, we will first determine the kinetic signatures of UV-induced DNA damage by SMRT DNA sequencing. In particular, we will derive the polymerase kinetic signatures for cyclobutane pyrimidine dimers (CPDs) and the pyrimidine (6-4) pyrimidone photoproducts (6-4) PPs formed by UV-radiation. In aim 2, we will use the kinetic signatures to directly detect CPDs and (6-4) PPs in UV-exposed melanocytes. The DDIP-enriched DNA fragments from the melanocytes will serve as templates for SMRT DNA sequencing of both strands. The kinetic analysis algorithms will be optimized to enable sensitive and accurate detection of the UV-DNA damage sites at the whole-genome level. Together, these aims will transform our understanding of how solar UV radiation, the most common human cancer etiological agent, leads to skin cancer and will facilitate new cancer-prevention strategies. Once developed, the DDIP-SMRT seq method can be extended to study the effects of other environmental carcinogens.